柳氮磺胺吡啶治疗冰醋酸诱导小鼠急性腹膜炎的作用机制

Mechanism of sulfasalazine in the treatment of acute peritonitis induced by glacial acetic acid in mice

目的应用冰醋酸诱导小鼠急性腹膜炎模型,探讨柳氮磺胺吡啶治疗小鼠急性腹膜炎的作用机制。方法60只C57BL/6雌性小鼠随机分为空白对照组、模型组和高、低剂量治疗组,每组15只。高、低剂量治疗组分别以200、100 mg/kg柳氮磺胺吡啶灌胃3 d后,高、低剂量治疗组和模型组使用体积分数1%冰醋酸腹腔注射诱导小鼠产生急性腹膜炎模型,空白对照组腹腔注射相同体积生理盐水;高、低剂量治疗组继续给予柳氮磺胺吡啶灌胃1周,模型组、空白对照组以相同体积生理盐水灌胃1周;记录分析各组小鼠疾病活动指数(disease activity index,DAI)。采用HE染色观察各组小鼠肠道结构变化并进行组织学评分。采用ELISA法检测各组小鼠肝脏、回肠组织谷胱甘肽(glutathione,GSH)、丙二醛(malonaldehyde,MDA)含量及血清肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素(interleukin-6,IL)-6和IL-1β水平;采用Western blot法检测各组小鼠回肠组织核转录因子-κB通路中相关蛋白p-p65、p65、p-IκB、IκB相对表达量。结果实验1~7 d,模型组、低剂量治疗组、高剂量治疗组小鼠DAI均高于空白对照组(P<0.05),低、高剂量治疗组小鼠DAI均低于模型组(P<0.05);实验2~7 d,高剂量治疗组小鼠DAI低于低剂量治疗组(P<0.05)。模型组小鼠肠黏膜损伤严重,高、低剂量治疗组小鼠肠道结构基本完整;模型组、低剂量治疗组、高剂量治疗组小鼠回肠组织组织学评分、肝脏和回肠组织MDA含量及血清TNF-α、IL-1β水平均高于空白对照组(P<0.05),模型组高于低、高剂量治疗组(P<0.05),低剂量治疗组高于高剂量治疗组(P<0.05);模型组、低剂量治疗组、高剂量治疗组肝脏和回肠组织GSH含量均低于空白对照组(P<0.05),模型组低于低、高剂量治疗组(P<0.05),低剂量治疗组低于高剂量治疗组(P<0.05);回肠组织p-p65、p65、p-IκB蛋白相对表达量在模型组(0.78±0.13、0.74±0.09、0.89±0.16)、低剂量治疗组(0.63±0.07、0.57±0.11、0.62±0.15)、高剂量治疗组(0.42±0.10、0.25±0.08、0.50±0.02)均高于空白对照组(0.35±0.04、0.19±0.05、0.45±0.03)(P<0.05),模型组高于低、高剂量治疗组(P<0.05),低剂量治疗组高于高剂量治疗组(P<0.05);回肠组织IκB蛋白相对表达量在模型组(0.28±0.04)、低剂量治疗组(0.49±0.06)、高剂量治疗组(0.65±0.02)均低于空白对照组(0.73±0.14)(P<0.05),模型组低于低、高剂量治疗组(P<0.05),低剂量治疗组低于高剂量治疗组(P<0.05)。结论柳氮磺胺吡啶可通过抑制核转录因子-κB信号通路的激活降低小鼠机体炎症水平,有效缓解小鼠急性腹膜炎的症状。

Objective To investigate the mechanism of salazamide pyridine in the treatment of acute peritonitis by using acute peritonitis mice models induced by glacial acetic acid.Methods Sixty female C57 BL/6 mice were randomly divided into blank control group,model group,high-dose treatment group and low-dose treatment group,with 15 mice in each group.High-and low-dose treatment groups received lavage with 200 and 100 mg/kg salazosul phapyridine for three days,respectively.The acute peritonitis mice models were prepared with intraperitoneal injection of 1%glacial acetic acid in high-dose treatment group,low-dose treatment group and model group,while blank control group was intraperitoneally injected with equivalent volume of normal saline.After modeling,high-and low-dose treatment groups received lavage with salazamide pyridine,while model group and blank control group received lavage with normal saline for one week.The disease activity index(DAI)was recorded and analyzed in each group.The changes in the intestinal structure were observed by HE staining and histopathological scoring was performed.ELISA was used to detect the contents of glutathione(GSH)and malonaldehyde(MDA)in the liver and ileum tissues,and the levels of tumor necrosis factor-α(TNF-α),interleukin(IL)-6 and IL-1β in the serum.Western blot was used to detect the expressions of p-p65,p65,p-IκB and IκB in nuclear factor-kappa B pathway in the ileum tissues.Results The DAI was higher in model group,low-dose treatment group and high-dose treatment group than that in blank control group(P<0.05),and higher in low-and high-dose treatment groups than that in model group from the first to the seventh day of experiment(P<0.05).The DAI was higher in high-dose treatment group than that in low-dose treatment group from the second to the seventh day of experiment(P<0.05).The intestinal mucosa was seriously damaged in model group,and the intestinal structure was basically intact in high-and low-dose groups.The histological score,MDA contents in the liver and ileum tissue,and the levels of TNF-α and IL-1β decreased gradually in turn in model group,low-dose treatment group,high-dose treatment group and blank control group(P<0.05).The contents of GSH in the liver and ileum increased gradually in turn in model group,low-dose treatment group,high-dose treatment group and blank control group(P<0.05).The relative expressions of p-p65,p65 and p-IκB in the ileum tissues decreased gradually in turn in model group(0.78±0.13,0.74±0.09,0.89±0.16),low-dose treatment group(0.63±0.07,0.57±0.11,0.62±0.15),high-dose treatment group(0.42±0.10,0.25±0.08,0.50±0.02)and blank control group(0.35±0.04,0.19±0.05,0.45±0.03)(P<0.05).The relative expression of IκB increased gradually in turn in model group(0.28±0.04),low-dose treatment group(0.49±0.06),high-dose treatment group(0.65±0.02)and blank control group(0.73±0.14)(P<0.05).Conclusion Salazosulfa pyridine can reduce the inflammation level by inhibiting the activation of NF-κB signaling pathway so as to relieve the symptoms of acute peritonitis effectively in mice.